Peptide for diagnosing and immunizing against T. cruzi infection

ABSTRACT

There is disclosed an antigenic peptide that comprises at least 15 amino acids having the sequence Ala Glu Pro Lys X Ala Glu Pro Lys X Ala Glu Pro Lys X, wherein X is Pro or Ser. This peptide is useful in an ELISA assay to detect antibodies specific to T. cruzi infection and Chagas disease. This peptide is further useful in a vaccine composition for immunizing an individual to prevent Chagas disease upon exposure to T. cruzi.

This invention was supported in part through grant number NIH AI22726from the National Institutes of Health. The U.S. Government, therefore,may have certain rights to this invention.

TECHNICAL FIELD OF THE INVENTION

The present invention relates to a 15 mer peptide that is the epitoperepeat sequence for a predominant antigen of T. cruzi. The inventivepeptide is useful for diagnosing T. cruzi infection and for use in avaccine to immunize an individual to reduce T. cruzi infection andclinical manifestations of Chagas disease.

BACKGROUND OF THE INVENTION

Chagas disease is one of the most important endemic problems in Centraland South America, for which no definitive chemotherapeutic orimmunological treatment is available. Trypanosoma cruzi (T. cruzi) isthe agent of Chagas disease. Infection with a protozoan parasite T.cruzi, the causitive agent of Chagas disease, occurs in an estimated 18million persons throughout Latin America and is a major cause of chronicheart disease. Immune responses after T. cruzi infection areparticularly complex due to the biochemical diversity of multipleparasite strains and influence of host-genetic factors. The result is awide diversity in clinical manifestations of Chagas disease and, in somecases, the disruption of immune regulation leading to immunosuppressionand/or development of autoimmunity. This parasite has a complex lifecycle involving an epimastigote stage in the insect vector and two mainstages in the mammalian host. One stage is present in blood(trypomastigote) and a second stage is intracellular (amastigote).

The acute phase of T. cruzi infection is often asymptomatic. Theinfection may remain quiescent for decades. Some patients may, however,develop a progressive chronic form of the disease with cardiac and/ordigestive tract alterations. After the acute phase with parasitemia,parasite growth is usually controlled by the host and patients oranimals enter into a chronic phase where few parasites are present inthe blood.

Immune responses to protozoan infection are complex, involving bothhumoral and cell-mediated responses to an array of parasite antigens.Infection often involves multiple life cycle stages of these parasites,which adds to the diversity of antigens potentially important for thedevelopment of protective immunity. To examine the molecular basis ofthe immune responses elicited during these infections, recent effortshave focused on evaluating responses to defined parasite B- and T-cellepitopes.

T. cruzi infections are often subtle and long-lasting, making diagnosiscrucial and problematic. Detecting antibodies against parasite antigensis a most common and reliable method of determining clinical andsubclinical infections. Presently, serological tests use whole or lysedT. cruzi and require positive results on two of three tests, includingcomplement fixation, indirect immunofluorescence, passive agglutination,or ELISA to accurately detect T. cruzi infection. The expense as well asdifficulty in performing such tests reliably prevent the screening ofblood or sera in many endemic areas.

Blood bank screening is particularly important in South America, where0.1-62% of samples may be infected and where the parasite is frequentlytransmitted by blood transfusion. It is also important and of increasingconcern that the blood supply in certain U.S. cities is contaminatedwith T. cruzi parasites.

Therefore, there is a need in the art for a greater understanding ofresponses to specific parasite antigens. Although several antigens of T.cruzi have been identified and characterized biochemically, limited dataare available on the evaluation of human immune responses to thesemolecules.

SUMMARY OF THE INVENTION

The present invention relates to the cloning and expression of a T.cruzi antigen gene sequence (SEQ ID NOs:1 and 2) encoding theimmunodominant protein with an essential repetitive epitope. This genesequence is conserved among diverse T. cruzi isolates. The inventiveantigenic peptide domain of T. cruzi is predominantly expressed bytrypomastigotes, the infective form of the parasite. Evaluation of humanimmune responses to this antigenic peptide domain of T. cruzi revealedeasily detectable levels of antibodies in greater than 95 percent of T.cruzi infected sera samples from several South American countries.

The antigenic peptide domain of T. cruzi comprises the amino acidsequence Ala Glu Pro Lys X₁ Ala Glu Pro Lys X₂ Ala Glu Pro Lys X₃, (SEQID NO:3) wherein X is Pro or Ser and when X₁ is Ser, X₃ is Ser, or whenX₁ is PrO, X₃ is Pro. The antigenic peptide can also comprise a linkersequence at either the N-terminus or the C-terminus of the antigenicpeptide domain wherein the linker sequence facilitates attachment orconjugation of the antigenic peptide domain to various carrier moleculesor solid support systems.

The present invention further comprises a method for diagnosing Chagasdisease or T. cruzi infection by detecting antibodies specific to theinventive antigenic peptide domain. This method comprises contacting asample of whole blood or an immunoglobulin-containing component of wholeblood with the inventive antigenic peptide conjugated to a solid phase,washing unbound antibodies from the solid phase, adding the inventiveantigenic peptide conjugated to a detectable moiety to form an antigenicpeptide-antibody complex, and detecting the antigenic peptide-antibodycomplex.

Further still, the present invention comprises a vaccine composition forimmunizing an individual for preventing Chagas disease symptoms of T.cruzi infection upon exposure to T. cruzi. The vaccine compositioncomprises an immunologically effective amount of the inventive antigenicpeptide and a vaccine adjuvant, such as Freund's adjuvant.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the DNA sequence and deduced amino acid sequence of a 636base pair TcD insert shown with residues blocked to indicate a 10-aminoacid repetitive unit and the number of repeats. Boxed amino acids markdegeneracies in the repeat unit. The DNA sequence and deduced amino acidsequence are also shown in SEQ ID NOs:1 and 2.

FIGS. 2A and 2B show an ELISA evaluation of recombinant TcD andsynthetic TcD peptide. Absorbance values are based upon a population of127 individuals with T. cruzi infection, 34 individuals withleishmaniasis, 10 with malaria, 17 mycobacterial infections and 32normal sera, against T. cruzi lysate (hatched bars) and recombinantantigenic peptide (double dash hatched bars). In FIG. 2B the adsorbancevalues are for 127 T. cruzi infection sera, 9 acute Chagas disease sera,15 other infected sera, including leishmaniasis, 10 malaria, 16mycobacterial infection and 32 normal sera. All sera samples wereevaluated with synthetic antigenic peptide.

DETAILED DESCRIPTION OF THE INVENTION

We have identified and synthesized a major antigenic epitope of anapproximately 260 kD T. cruzi antigen expressed predominantly bytrypomastigotes. This antigenic peptide domain of T. cruzi is conservedamong geographically diverse T. cruzi isolates. Conservation of theantigen was further indicated by the presence of TcD-specific antibodiesin sera from Chagas patients having great clinical and geographicaldiversity, produced as a result of natural infection with T. cruziparasites expressing the antigenic peptide repetitive epitope. Thisantigenic peptide domain of T. cruzi is useful for diagnosing T. cruziinfection, Chagas disease, and for use in a vaccine composition toprotect individuals from Chagas disease or other lethal complicationsupon exposure to T. cruzi parasites.

Response to the antigenic peptide domain of T. cruzi was found to havesuch extremely high prevalence in seroreactive Chagas patients that ithas usefulness as a diagnostic agent. Particularly, the antigenicpeptide domain induces antibodies in such patients such that infectionby T. cruzi or symptoms of Chagas disease can diagnosed by the presenceof such antibodies using an ELISA-RW diagnostic assay with recombinantor synthetic antigenic peptide from T. cruzi.

The antigenic peptide domain of T. cruzi comprises the protein sequenceAla Glu Pro Lys X₁ Ala Glu Pro Lys X₂ Ala Glu Pro Lys X₃, (SEQ ID NO:3)wherein X is Pro or Ser and when X₁ is Ser, X₃ is Ser, or when X₁ isPrO, X₃ is PrO. The antigenic peptide comprises at least a 15 residuesequence having three groups of five amino acids as a repeat sequence.The antigenic peptide domain can have additional five residue Ala GluPro Lys X (SEQ ID NO:4) sequences to cause the antigenic peptide to be15, 20, 25, 30, 35, 40, etc., amino acids in length.

Moreover, mice immunized with the inventive antigenic peptide have beenprotected from lethal T. cruzi infection. Therefore, the inventiveantigenic peptide can be used as a vaccine to prevent generalizedinfection from T. cruzi and enhance tile host immune response to T.cruzi exposure.

The present inventive antigenic peptide is the first such antigen havingan epitope with serodiagnostic potential. A previously reported clonedT. cruzi antigen (Ibanez et al., Mol. Biochemical Parasitology 25:175,1987; Ibanez et al., Mol. Biochemical Parasitology 30:27, 1988; andAffranchino et al., Mol. Biochemical Parasitology 34:221, 1989)contained repetitive domains, with one domain reportedly present in an85 kD antigen. Although the 5-amino acid repeat sequence in the 85 kDantigen is similar to the second half of the repeat sequence of theinventive antigenic peptide domain, reactivity with the 85 kD antigendemonstrated only 40% positives in Chagas patient sera. The presentinventive peptide, by contrast, exhibited confirmation rates of greaterthan 95%. This result is consistent with our mapping studies, whichindicated that peptides containing only the 5-residue repeat sequencelacked an essential portion of a dominant B-cell epitope of TcD.

In ELISA assay with the synthetic antigenic peptide domain according tothe present invention is easy to perform, allows for standardization ofreagents, permits screening of large numbers of samples, and can be usedwith either blood or serum samples.

We purified a series of T. cruzi antigens to find an antigenic peptidedomain of a T. cruzi antigen responsible for uniform epitope binding ina vast majority of T. cruzi infected individuals. We made a genomicexpression library in λZAPII with mechanically sheared DNA of T. cruzi.Recombinants expressing T. cruzi antigen genes were selected based upontheir reactivity with a pool of Chagas patients' sera, preadsorbed toremove anti-E. coli reactivity. Of twelve clones identified, one clone,called TcD, was exceptionally reactive with the pooled patients' sera.Purified recombinant antigen of clone TcD migrated at about 59 kD onSDS/PAGE. In an immunoblot analysis, the TcD antigen was stronglyrecognized by pooled Chagas patient sera but not recognized by a pool ofnormal sera obtained from normal volunteers in Seattle, Wash. Moreover,a pool of high-titer sera from patients with acute visceralleishmaniasis, an infection known to induce antibodies cross-reactivewith T. cruzi, was negative for the TcD antigen.

The sequence of the TcD antigen is shown in FIG. 1. Clone TcD encodes a10-amino acid repetitive sequence. DNA sequence analysis of clone TcDpredicted an amino acid sequence comprised entirely of a 10-amino acidrepeat sequence arrayed in tandem, and present in 20.5 copies with minordegeneracies in 5 positions (FIG. 1). The predicted molecular weight ofrecombinant, unglycosylated TcD antigen was 36.3 kD. The migration ofthe TcD antigen at 59 kD observed during SDS/PAGE most likely reflecteda high proline content (28%).

A 636 base pair insert of clone TcD was used to probe Southern blots ofT. cruzi DNA and DNA from several other protozoan parasites of humans.The probe hybridized to multiple restriction fragments of T. cruzi DNAbut not to the other protozoan parasites including T. brucei, Leishmaniachagasi, L. amazonensis, L. donovani, and T. rangeli. Analysis of DNAfrom seven geographically diverse T. cruzi isolates indicate that TcDgene sequence was conserved among isolates showing restriction fragmentlink polymorphism and variability in gene-copy number.

The antigenic peptide of T. cruzi comprises at least 15 amino acidshaving the sequence Ala Glu Pro Lys X Ala Glu Pro X Ala Glu Pro Lys X,wherein X is Pro or Ser. Additional 5 amino acid sequences (Ala Glu ProLys X) may be added to the basic 15 residue antigenic peptide domain ofT. cruzi. A further sequence may be added to the antigenic peptides tolink this peptide at either its N-terminal or C-terminal wherein thelinker sequence facilitates attachment or conjugation of the antigenicpeptide to carrier molecules. An example of a linker sequence is Gly CysGly. The antigenic peptide is made, preferably, by synthetic means on aprogrammable peptide synthesizer.

ELISA assays have been conducted utilizing antigens or antibodies as theouter components of a sandwich. An ELISA assay of blood or sera fromindividuals can detect T. cruzi infection of Chagas disease by anantibody specific to the antigenic peptide domain. Therefore, onecomponent of an ELISA sandwich comprises the antigenic peptide of thepresent invention. Another component comprises an agent that can bind tothe anti-T. cruzi antibody include, for example, anti-immunoglobulin orprotein A. Each component can form a antigenic peptide-antibody complexwhich contains a detectable moiety. The detectable moiety is known inthe art of ELISA diagnostic assays as that component that identifies theantigenic peptide-antibody complex through visual, fluorescent,radionuclide or other means. Common examples of detectable moietiesinclude fluorescent or chemiluminscent agents or enzymes such ashorseradish peroxidase.

In a series of studies, patient sera from T. cruzi infected individualsor Chagas patients was compared with sera from patients infected withother parasites or normal sera. Patient sera with ELISA values at leastfive standard deviations greater than mean adsorbance value of normalcontrols were considered positive. Of confirmed T. cruzi infected sera,greater than 95 percent (121 of 127) were positive for an anti-TcDantibody. Therefore, detection of an anti-TcD antibody in T. cruziinfected individuals is a reliable method of detecting Chagas disease orT. cruzi infection.

EXAMPLE 1

This example illustrates cloning of the TcD antigen from T. cruzi. Agenomic library was constructed in λZAPII (Stratagene) with mechanicallysheared DNA of T. cruzi. Construction of the library and excision of apBFK (-) phagemid sequences were performed according to manufacturer'sprotocols. Recombinants were screened with a pool of Chagas patients'sera preadsorbed to remove anti-E. coli reactivity as described inSambrook et al., Molecular Cloning: A Laboratory Manual, Cold SpringHarbor Laboratories, Cold Spring Harbor, N.Y., 1989. A 59 kD recombinantantigen, called clone TcD, was purified from a soluble lysate of inducedbacterial cultures by ammonium sulfate fractionation, preparativeisoelectric focusing with a Bio-Rad Rotofor IEF cell and 1%-3/10ampholytes in the presence of 8M urea followed by SDS/PAGE andelectroelution as described in Reed et al., J. Clin. Invest. 85:690,1990. Protein concentrations were determined using a Pierce BCA proteinassay.

Patients' sera were collected from well-characterized patients withacute or chronic Chagas disease or indeterminant T. cruzi infection fromthe South American countries Brazil (Northern and Southern), Bolivia andArgentina. Normal sera were obtained from individuals living innon-endemic T. cruzi areas (Seattle). Sera from confirmed visceral orcutaneous leishmaniasis were obtained from parasitologically confirmedSudanese patients. Mycobacterial infection sera were obtained fromSeattle (tuberculosis) or Haiti for leprosy.

Twelve clones were identified. One clone, called TcD, was exceptionallyreactive with patients' sera. Purified recombinant antigen from cloneTcD migrated at 59 kD on SDS/PAGE. An immunoblot analysis found that theTcD antigen was strongly recognized by Chagas patients' serum but notrecognized by normal sera or by high-titer sera from patients with acutevisceral leishmaniasis, an infection known to induce antibodiescross-reactive with T. cruzi.

EXAMPLE 2

This example illustrates an evaluation of synthetic antigenic peptidesderived from the antigenic peptide domain of T. cruzi having amino acidlength of 5, 10, 15, and 20 amino acids in length. The syntheticpeptides were constructed with five amino acid repeat sequences of AlaGlu Pro Lys X, wherein X is Pro or Ser. The data (FIG. 2) show that apeptide containing 15 residues of the repeat sequence was required tomap the immunodominant B-cell epitope of TcD. Moreover, the 15 aminoacid synthetic peptide had reactivity of patient sera comparable to thatobtained with the recombinant molecule. One hundred sixteen of onehundred twenty Chagas sera patients gave positive adsorbance values. Inaddition, positive TcD-specific antibody responses were detected in 8 of9 acute Chagas disease patients, indicative of an early immune responseto T. cruzi infection to this epitope.

EXAMPLE 3

This example illustrates the usefulness of the inventive antigenicpeptide in a vaccine composition to reduce complication and mortalityassociated with T. cruzi infection. A group of 8 week old female C57/6mice (Jackson Labs, Bar Harbor, Me.) were divided into three treatmentgroups. Group A (5 mice) was the control group and received no vaccinetreatment. Group B was a second control group of 4 mice that receivedonly adjuvant treatment. Group C (4 mice) received the inventive vaccinecomposition comprising the antigenic peptide (TcD) (15 mer) plus anadjuvant.

The treatment schedule was vaccine administration (s.c.) on day 0,wherein the vaccine comprised 200 μg TcD peptide in complete Freund'sadjuvant. Vaccine was administered (i.p.) on day 24 comprising 100 μgTcD. Vaccine was also administered (s.c.) on day 50 wherein this vaccinecomposition comprised 200 μg TcD peptide in incomplete Freund's adjuvantplus 25 μg muramyl dipeptide (MDP, Calbiochem) an additional adjuvantagent.

At day 57, each mouse was challenged with 1,000 T. cruzi (TcTc²) ip. Atday 75 (or 18 days post challenge) peak parasitemia was determined andeach animal was observed for mortality. The data presented in Table 1below show the protective effects of the TcD peptide vaccine compositionto T. cruzi challenge.

                  TABLE 1                                                         ______________________________________                                        Group  Treatment     Peak Parasitemia                                                                            Mortality                                  ______________________________________                                        A      nothing       4 × 10.sup.5 ± 5 × 10.sup.5                                                  5/5                                        B      adjuvant      4 × 10.sup.5 ± 5 × 10.sup.5                                                  4/4                                        C      TcD + adjuvant                                                                              1 × 10.sup.5 ± 8 × 10.sup.4                                                  1/4                                        ______________________________________                                    

    __________________________________________________________________________    SEQUENCE LISTING                                                              (1) GENERAL INFORMATION:                                                      (iii) NUMBER OF SEQUENCES: 4                                                  (2) INFORMATION FOR SEQ ID NO:1:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 636 base pairs                                                    (B) TYPE: nucleic acid                                                        (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: cDNA to mRNA                                              (iii) HYPOTHETICAL: NO                                                        (iv) ANTI-SENSE: NO                                                           (vi) ORIGINAL SOURCE:                                                         (A) ORGANISM: Trypanosoma cruzi                                               (vii) IMMEDIATE SOURCE:                                                       (B) CLONE: TcD                                                                (ix) FEATURE:                                                                 (A) NAME/KEY: CDS                                                             (B) LOCATION: 8..628                                                          (xi) SEQUENCE DESCRIPTION: SEQ ID NO:1:                                       GAATTCAGCAGAGCCCAAACCAGCGGAGCCGAAGTCAGCAGAGCCTAAA49                           AlaGluProLysProAlaGluProLysSerAlaGluProLys                                    1510                                                                          CCAGCGGAGCCGAAATCGGCAGAGCCCAAACCAGCGGAGCCGAAATCG97                            ProAlaG luProLysSerAlaGluProLysProAlaGluProLysSer                             15202530                                                                      GCAGAGCCCAAACCAGCGGAGCCGAAATCAGCGGGGCCTAAACCAGCG145                           Ala GluProLysProAlaGluProLysSerAlaGlyProLysProAla                             354045                                                                        GAGCCGAAGTCAGCGGAGCCTAAACCAGCGGAGCCGAAATCAGCAGAG193                           Glu ProLysSerAlaGluProLysProAlaGluProLysSerAlaGlu                             505560                                                                        CCCAAACCAGCGGAGCCGAAATCGGCAGAGCCCAAACCAGCGGAGCCG241                           ProLy -ProAlaGluProLysSerAlaGluProLysProAlaGluPro                             657075                                                                        AAGTCAGCAGAGCCCAAACCAGCGGAGTCGAAGTCAGCAGAGCCTAAA289                           LysSerAlaG luProLysProAlaGluSerLysSerAlaGluProLys                             808590                                                                        CCAGCGGAGCCGAAATCAGCAGAGCCCAAACCAGCGGAGTCGAAGTCA337                           ProAlaGluProLysSer AlaGluProLysProAlaGluSerLysSer                             95100105110                                                                   GCAGAGCCCAAACCAGCGGAGCCGAAGTCAGCAGAGCCCAAACCAGCG385                           AlaGluProLysPro AlaGluProLysSerAlaGluProLysProAla                             115120125                                                                     GAGCCGAAGTCAGCAGAGCCCAAACCAGCGGAGCCGAAATCAGCGGAG433                           GluProLysSerAl aGluProLysProAlaGluProLysSerAlaGlu                             130135140                                                                     CCCAAACCAGCGGAGCCGAAATCAGCAGAGCCCAAACCAGCGGAGTCG481                           ProLysProAlaGluP roLysSerAlaGluProLysProAlaGluSer                             145150155                                                                     AAATCAGCGGGGCCTAAACCAGCGGAGCCGAAGTCAGCGGAGCCAAAA529                           LysSerAlaGlyProLysPro AlaGluProLysSerAlaGluProLys                             160165170                                                                     CCAGCGGAGCCGAAATCAGCGGAGCCAAAACCAGCGGAGCCGAAATCG577                           ProAlaGluProLysSerAlaGluProLys ProAlaGluProLysSer                             175180185190                                                                  GCAGAGCCCAAACCAGCGGAGCCGAAGTCAGCAGAGCCAAAACCAGCG625                           AlaGluProLysProAlaGluProLy sSerAlaGluProLysProAla                             195200205                                                                     GAGCCGAATTC636                                                                Glu                                                                           (2) INFORMATION FOR SEQ ID NO:2:                                              (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 207 amino acids                                                  (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2:                                       AlaGluProLysProAlaGluProLysSerAlaGluProLysProAla                              1510 15                                                                       GluProLysSerAlaGluProLysProAlaGluProLysSerAlaGlu                              202530                                                                        ProLysProAlaGluProLysSerAlaGlyProLysProAlaGluPro                               354045                                                                       LysSerAlaGluProLysProAlaGluProLysSerAlaGluProLys                              505560                                                                        ProAlaGluProLysSerAlaGluPr oLysProAlaGluProLysSer                             65707580                                                                      AlaGluProLysProAlaGluSerLysSerAlaGluProLysProAla                              859 095                                                                       GluProLysSerAlaGluProLysProAlaGluSerLysSerAlaGlu                              100105110                                                                     ProLysProAlaGluProLysSerAlaGluProLysProA laGluPro                             115120125                                                                     LysSerAlaGluProLysProAlaGluProLysSerAlaGluProLys                              130135140                                                                     ProAlaGluProLys SerAlaGluProLysProAlaGluSerLysSer                             145150155160                                                                  AlaGlyProLysProAlaGluProLysSerAlaGluProLysProAla                              165 170175                                                                    GluProLysSerAlaGluProLysProAlaGluProLysSerAlaGlu                              180185190                                                                     ProLysProAlaGluProLysSerAlaGl uProLysProAlaGlu                                195200205                                                                     (2) INFORMATION FOR SEQ ID NO:3:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 15 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (iii) HYPOTHETICAL: NO                                                         (iv) ANTI-SENSE: NO                                                          (ix) FEATURE:                                                                 (1) OTHER INFORMATION: Xaa is Pro or Ser; when Xaa in                         position 5 is Ser, Xaa in position 10 is Pro and Xaa in                       position 15 is Ser; when Xaa in position 5 is Pro, Xaa in                     position 10 is Ser and Xaa in position 15 is Pro.                             (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3:                                       AlaGluProLysXaaAlaGluProLys XaaAlaGluProLysXaa                                151015                                                                        (2) INFORMATION FOR SEQ ID NO:4:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 5 amino acids                                                     (B) TYPE: amino acid                                                          (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                    (iii) HYPOTHETICAL: NO                                                       (iv) ANTI-SENSE: NO                                                           (ix) FEATURE:                                                                 (1) OTHER INFORMATION: Xaa is Pro or Ser.                                     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4:                                       AlaGluProLysXaa                                                               15                                                                        

I claim:
 1. A major antigenic epitope of a 260 Kd T. cruzi antigen,comprising the amino acid sequence Ala Glu Pro Lys Xaa Ala Glu Pro LysXaa Ala Glu Pro Lys Xaa (SEQ ID NO:1), wherein Xaa is Pro or Ser andwhen Xaa in position 5 is Ser, Xaa in position 10 is Pro and Xaa inposition 15 is Ser, or when Xaa in position 5 is Pro, Xaa in position 10is Ser and Xaa in position 15 is Pro.
 2. The peptide of claim 1 furthercomprising one or a plurality of Ala Glu Pro Lys Xaa (SEQ ID NO:4)peptide sequences wherein Xaa is Pro or Ser.
 3. The peptide of claim 1further comprising a linker sequence at either the N-terminal or theC-terminal, wherein the linker sequence facilitates attachment orconjugation of the antigenic peptide to carrier molecules.
 4. Apharmaceutical composition for prophylactic administration to anindividual to reduce complication and mortality associated with T. cruziinfection, comprising an immunologically effective amount of theantigenic peptide of claim 1 and a vaccine adjuvant.
 5. Thepharmaceutical composition of claim 4 wherein the vaccine adjuvant is aFreund's adjuvant.